Fundamental aspects of the neuroendocrine, autonomic and behavioral responses to stress are integrated by parvocellular corticotropin releasing hormone (CRH) neurons in the hypothalamic paraventricular nucleus (PVN). The main goal of the proposed experiments is to elucidate the role of the neuropeptide pituitary adenylate cyclase-activating polypeptide (PACAP) in the central activation of parvocellular CRH neurons and to integrate the PACAPergic mechanisms into specific neuroanatomical pathways subserving the stress response. Based on our preliminary data, we hypothesize that PACAP is an endogenous activator of parvocellular CRH neurons and its actions are exerted by PACAP containing innervation, with the involvement of intracellular signaling pathways that activate the transcription factor CREB by phosphorylation. Aim 1, using in vivo pharmacologic approaches with local intra-PVN microinjections, will investigate the role of PACAP in transcriptional activation of CRH neurons as well as its interactions with adrenergic mechanisms. Aim 2 will determine the significance of endogenous PACAP in the PVN for the activation of CRH neurons, following systemic and psychological stressors, using selective PACAP receptor antagonist microinjections into the PVN. Aim 3 will identify the brain region-specific contribution of PACAP innervation to CRH neurons in the PVN from neuroanatomical tract-tracing experiments combined with multiple-labeling immunofluorescence. Aim 4 will directly test the hypothesis whether PACAP neurons are activated by stress in brain regions that send innervation to the PVN. Thus, combined results from these aims will reveal the physiological role of PACAP for the central neuroendocrine effector system of the hypothalamic-pituitary-adrenal axis in acute stress. Synthesis of information from the proposed experiments may become applicable toward a better understanding of the pathophysiology of certain stress-related illnesses.